Molecule separation device and method combining multiple filtration media

ABSTRACT

A molecule separator device for isolating molecules having at least two separable properties and within a solution. The device includes a housing, and at least two molecule collection media disposed within the housing, whereby each such medium captures molecules exhibiting a respective property. In one embodiment, a first membrane captures only molecules with an ionic and/or hydrophobic and/or affinity attraction property while a second membrane captures only such molecules that additionally fall within a particular molecular weight range. A preferred housing is cylindrical for acceptance within a centrifuge, and is constructed of a plurality of releasably-connected compartments. The collection media is sequentially situated and centrifugation of the housing drives the solution through the media. Because of separation and subsequent collection in one device of molecules bearing multiple properties, the present invention permits rapid and efficient isolation of molecules and micro-particulate having a plurality of identification characteristics.

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefits of U.S. Provisional PatentApplication Ser. No. 60/193,118 filed Mar. 30, 2000, and of U.S.Provisional Patent Application Ser. No. 60/198,529 filed Apr. 20, 2000.

STATEMENT RE: FEDERALLY SPONSORED RESEARCH/DEVELOPMENT

[0002] Not Applicable

[0003] BACKGROUND OF THE INVENTION

[0004] This invention relates in general to the separation and captureof molecule types from a solution mixture thereof, and in particular toapparatus and methodology wherein molecules with two or more definedproperties such as ionic, hydrophobic, or affinity attractions andmolecular weight ranges are captured and retained first for one suchproperty and thereafter for the additional property, with suchrespective collections accomplished sequentially in a single moleculeseparator device.

[0005] One of the most important tasks performed during research andother laboratory procedures is the separation of certain components froma mixture of components such that chemical or other analysis canproceed. A usual manner of accomplishing such separations is theemployment of filtration devices whereby filtrate is collected by afilter medium as a solution containing the filtrate product passesthrough the filter medium. The most common of filter media are filtermembranes and matrices thereof whose interstices prohibit, and thuscapture, particulate whose physical size is too large to pass through aspart of the solute.

[0006] While such filter membranes and related matrices (e.g. cloth)work well where particulate to be collected is defined only according tosize and the interstices of the filter medium are adequately sized forfiltrate retention, the separation of smaller particulate, asexemplified at the molecular level, requires much greater sophisticationin order to accomplish separation and collection. Additionally,molecular separation many times involves the need to collect moleculesthat must possess at least two properties such as ionic, hydrophobic, oraffinity attractions plus a limited molecular weight range. Toaccomplish separation and collection of such micro-particulate, multiplefiltration devices must be employed where each device has aone-membrane-type filter for collecting filtrate having one definedcharacteristic from a solution. Once molecules are collected thatpossess the first desired property, the filtrate must be transferred toa second filtration device having a second one-membrane-type filter thataddresses the second property and collects molecular filtrate meetingthe second standard.

[0007] As is thus apparent, where, for example, molecules having atleast two defining characteristics are to be isolated from a solution, auser must inefficiently perform filter procedures at least two separatetimes using at least two separate filtration devices. In view of thisnow-required inefficient approach, it is a primary object of the presentinvention to provide a molecule separator device where molecules havinga plurality of properties can be separated and collected with oneseparator device.

[0008] Another object of the present invention to provide a moleculeseparator device where such molecule separation is accomplishedsequentially within a single housing.

[0009] Yet another object of the present invention to provide a moleculeseparator device where respective dedicated membrane media providefiltrate collection.

[0010] Still another object of the present invention is to providemethodology for separating and capturing molecules having a plurality ofproperties utilizing a single separator device.

[0011] These and other objects of the present invention will becomeapparent throughout the description thereof which now follows.

BRIEF SUMMARY OF THE INVENTION

[0012] The present invention is a molecule separator device forseparating and isolating molecules having at least two separableproperties and present in a solution comprising the molecules. Theseparator device includes a housing for accepting pressured passagethere through of the solution, and at least two molecule collectionmedia disposed within the housing, wherein each such medium capturesmolecules exhibiting a respective property respectively capturable bythe media. In a preferred embodiment, a first molecule-collectionchromatography membrane captures and retains only molecules with anionic, hydrophobic, or affinity attraction property while a secondmolecule-collection ultrafiltration membrane captures and retainsadditional such molecules that additionally fall within a particularmolecular weight range. Conversely, these exemplary membranes can be inreverse order such that the first molecular collection membrane is anultrafiltration membrane while the second membrane possesses the ionic,hydrophobic, or affinity attraction property. A preferred housing isgenerally cylindrical for operational acceptance within a generallycylindrical fixed-angle or swinging-bucket chamber of a centrifuge head,and is constructed of a plurality of liquid-tight, releasably-connectedcompartments in communication with each other. The collection media issituated in a sequential relationship among the compartments whilecentrifugation of the housing drives the solution through the media.Removing and replacing appropriate compartments during the moleculecollection process permits separate and replaceable reservoir, wash, andcollection sites to yield filtrate product as so chosen for furtheranalysis, processing, or use, or for discard where a separation goal isthe provision of clean solute. Because of separation and subsequentcollection of molecules bearing two or more properties, the presentinvention permits rapid and efficient isolation of molecules and/ormicro-particulate having multiple identification characteristics.

BRIEF SUMMARY OF THE DRAWINGS

[0013] An illustrative and presently preferred embodiment of theinvention is shown in the accompanying drawings in which:

[0014]FIG. 1 is a perspective view of a first embodiment of a moleculeseparator device for capture or collection of molecules and/ormicro-particulate;

[0015]FIG. 2 is a perspective view of a separated compartment structurefor the separator device of FIG. 1;

[0016]FIGS. 3a-3 e illustrate use of the embodiment of FIG. 1;

[0017]FIG. 4 is a side perspective view of a second embodiment of amolecule separator device; and

[0018]FIG. 5a-5 g illustrate use of the embodiment of FIG. 4.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0019] Referring first to FIGS. 1 and 2, a molecule separator device 10is shown. The device 10 includes a housing 12 constructed of tworeleasably connected, liquid-tight, separable compartments 14, 16attached to each other by conventional friction fit between adjacentcompartments. Within the housing 12 are two sequentially disposedmembranes 24, 26 for collecting filtrates. In particular, the firstmembrane 24 is a chromatography membrane operating as a cationic oranionic ion-exchange membrane, hydrophobic membrane, affinity membrane,or a combination thereof for attracting molecules exhibiting ionicand/or hydrophobic and/or affinity attractions. The first membrane 24can have a porosity non-limitedly exemplified in the range of 0.1 to 10microns and is fabricated of any appropriate microporous materialincluding nylon, polycarbonate, polyethersulfone, glass fiber,polypropylene, polysulfone, cellulose acetate, regenerated cellulose,and mixed esters of cellulose or other polymeric material as would berecognized by a skilled artisan. The second membrane 26 preferably isanisotropic (asymmetrical) and can be fabricated of the same materialsas the first while providing ultrafiltration in speaking towardmolecular weight characteristics for capturing molecule filtrate. Thus,a chosen molecular weight range can be exemplified in values from about5×10² to about 3×10⁶ Daltons.

[0020] As shown in FIG. 1, the upper compartment 14 of the housing 12has an upper reservoir chamber 28 immediately above the first membrane24 and a lower reservoir chamber 30 immediately below the first membrane24. The lower compartment 16 includes an upper chamber 32 immediatelyabove the second membrane 26 and a fluid collection chamber 34immediately beneath the second membrane 26. FIG. 2 shows an independentcompartment 36 attachable to the upper compartment 14 during certainwashing procedures as described later. The housing 12 can be constructedof a semi-rigid material such as polypropylene or of any other plasticor polymeric material as would be evident to a skilled artisan.Likewise, housing size can be as required to provide volumetricaccommodations as required for a particular task. A screw-type closurecap 38 with an aperture 40 there through closes the housing 12. As isapparent, the housing 12 resembles the configuration of a standardcentrifuge tube, thus permitting placement of the separator device 10within a standard fixed-angle or swinging-bucket chamber (not shown) ofa centrifuge head (not shown). While centrifugation is the preferredmanner of pressurized force, the aperture 40 in the screw cap 38 isprovided to accept a pressure nozzle such as the outlet of a hypodermicsyringe (not shown) whose pressure can be applied to force the solutionthrough the separator device 10.

[0021] A description of an exemplary operation of the separator device10 is accompanied by the illustrations of FIGS. 3a-3 e. First, the uppercompartment 14 and an independent compartment 36 are attached as shownin FIG. 3a. A subject solution is placed within the upper reservoirchamber 28 of upper the compartment 14, the cap 38 is secured in placeas shown in FIG. 3b, and the resulting unit is centrifuged (fixed angleor swinging bucket) or pressurized for as long as necessary (many timesabout 0.5 minute) to accomplish liquid movement through the unit. Asexpected, the force moves the liquid quickly through the first membrane24 as target molecules are collected. Since this first membrane 24 has arelatively large pore size, virtually any sized molecules ormicro-particulate can pass through unimpededly, and only targetmolecules or micro particulate with ionic, hydrophobic, or affinityattractions will be retained. Alternatively, dependent upon theproperties of the passing solution, target molecules ormicro-particulate may pass through the membrane while contaminant isretained. The cap 38 is removed, an appropriate buffer solution is addedto the upper compartment 14 which is re-capped, and a second period ofcentrifugation or pressurization is completed to assure removal of anycontaminants from the target molecules, while the molecules ormicro-particulate remain bound to the first membrane 24. Elution oftarget molecules is accomplished as the independent compartment 36 withsolute therein is removed and replaced with the lower compartment 16 asshown in FIG. 3c. The upper reservoir chamber 28 is then filled with anappropriate elution buffer to remove the target molecules from the firstmembrane 24 and the separator device 10 is centrifuged for severalminutes as the target molecules now pass through the first membrane 24are captured because of size by the second membrane 26. The uppercompartment 14 (FIG. 3d) is removed and, thereafter, the upper reservoirchamber 15 is filled with a final washing buffer and centrifuged forseveral minutes for product desalting and placing the target moleculesin a desired buffer such as physiological saline. Finally, anindependent compartment 36 (FIG. 3e) is placed onto the compartment 16,and the resulting unit is inverted and centrifuged or pressurized forfinal product collection as the target molecules are forced from thesecond membrane 26 and into the independent compartment 36.

[0022]FIGS. 4 and 5a-5 g show a second preferred embodiment and use of amolecule or micro-particulate separator device 50. In particular, theseparator device 50 includes a housing 52 constructed of two releasablyconnected, liquid-tight, separable compartments 54, 56, each having oneseparable reservoir 53, 57, with compartments 54, 56 and reservoirs 53,57 held to each adjacent structure by conventional friction fit. Withinthe housing 52 are two sequentially disposed membranes 63, 65 forcollecting two different filtrates. In particular, the first membrane 63is anisotropic (asymmetrical) and can be fabricated of any appropriatepolymeric material with ultrafiltration pore size including nylon,polycarbonate, polyethersulfone, glass fiber, polypropylene,polysulfone, cellulose acetate, regenerated cellulose, and mixed estersof cellulose or polymeric materials as would be recognized by a skilledartisan while providing ultrafiltration in speaking toward molecularweight characteristics for capturing molecule filtrate. Thus, a chosenmolecular weight range can be exemplified in values from about 5×10² toabout 3×10⁶ Daltons. The second membrane 65 is a chromatography membraneoperating as a cationic or anionic ion-exchange membrane, hydrophobicmembrane, affinity membrane, or a combination thereof for attractingmolecules exhibiting ionic and/or hydrophobic and/or affinityattractions. The second membrane 65 can have a porosity non-limitedlyexemplified in the range of 0.1 to 10 microns and is also fabricated ofnylon, polycarbonate, polyethersulfone, polysulfone, cellulose acetate,glass fiber, polypropylene, regenerated cellulose, and mixed esters ofcellulose or other polymeric materials.

[0023] As shown in FIG. 4, the upper compartment 54 of the housing 52has an upper reservoir chamber 58 immediately above the first membrane63 and a lower reservoir chamber 60 immediately below the first membrane63. The lower compartment 56 includes an upper chamber 62 immediatelyabove the second membrane 65 and a fluid collection chamber 64immediately beneath the second membrane 65. The housing 52 can beconstructed of a semi-rigid material such as polypropylene or of anyother polymeric material as would be evident to a skilled artisan.Likewise, housing size can be as required to provide volumetricaccommodations as required for a particular task. As is apparent, thehousing 52 resembles the configuration of a standard centrifuge tube,thus permitting placement of the separator device 50 within a standardfixed-angle or swinging-bucket chamber (not shown) of a centrifuge head(not shown).

[0024] A description of an exemplary operation of the separator device50 is accompanied by the illustrations of FIGS. 5a-5 g. First, a subjectsolution is placed within the upper chamber 62 of the lower compartment56 (FIG. 5a), the upper and lower compartments 54, 56 are attached asshown in FIG. 5b, and the resulting unit is centrifuged (fixed angle orswinging bucket) for as long as necessary (many times about 0.5 minute)to accomplish liquid movement through the membrane. As expected, thecentrifugal force moves the liquid quickly through the second membrane65 as target molecules are collected. Since this second membrane 65 hasa relatively large pore size, virtually any sized molecule ormicro-particulate can pass through unimpededly, and only targetmolecules with ionic or hydrophobic or affinity attractions will beretained. Alternatively, dependent upon the properties of the passingsolution, target molecules or micro-particulate may pass through themembrane while contaminant is retained. Next, an appropriate buffersolution is added to the upper chamber 62 of the lower compartment 56,and a second centrifugation is completed to assure removal of anycontaminants from the target molecules while the molecules remain boundto the second membrane 65. The reservoir 57 is then removed and emptied,and filled with an elution buffer. Upon reassembly, the separator device50 is inverted (FIG. 5e) and inserted into the centrifuge forcentrifugation to remove the target molecules or micro-particulate fromthe second membrane 65 and capture them because of size at the firstmembrane 63. Thereafter, while remaining in the now-upside downposition, the lower reservoir chamber 60 is filled with an appropriatebuffer to wash the target molecules free of high salt of the elutionbuffer while retaining the molecules at the first membrane 54. Finally,the reservoir 53 is emptied (FIG. 5f), the reservoir 57 is removed andreplaced with a new reservoir 57 a (FIG. 5g), and the resulting unit isinverted and centrifuged for final product collection as the targetmolecules are forced into the reservoir 57 a. Alternatively, of course,the device 50 may be inverted at the beginning of the process such thatthe ultrafiltration membrane is the first contact membrane.

[0025] As is apparent, the molecule separator devices above describedprovide rapid two-stage separations within a single, convenient, andmolecular-property specific apparatus. Additionally, as recognized bythe skilled artisan, there are numerous possible combinations ofchromatography membranes and ultrafiltration membranes for producingunique purification results. Therefore, while an illustrative andpresently preferred embodiment of the invention has been described indetail herein, it is to be understood that the inventive concepts may beotherwise variously embodied and employed and that the appended claimsare intended to be construed to include such variations except insofaras limited by prior art.

What is claimed is:
 1. A molecule separator device for separating andisolating molecules having at least two separable properties and presentin a solution comprising said molecules and/or micro-particulate, theseparator device comprising: a) a housing for accepting pressuredpassage there through of the solution; and b) at least two moleculecollection media disposed within the housing, wherein each said mediumcaptures molecules exhibiting a respective property respectivelycapturable by said media.
 2. A molecule separator device as claimed inclaim 1 wherein the molecule-collection media are membranes.
 3. Amolecule separator device as claimed in claim 1 wherein a first saidmedium captures molecules exhibiting a property of ionic and/orhydrophobic and/or affinity attraction.
 4. A molecule separator deviceas claimed in claim 3 wherein a second said medium captures moleculesexhibiting a molecular weight property from about 5×10² to about 3×10⁶Daltons.
 5. A molecule separator device as claimed in claim 1 wherein asecond said medium captures molecules exhibiting a molecular weightproperty from about 5×10² to about 3×10⁶ Daltons.
 6. A moleculeseparator device as claimed in claim 1 wherein the at least twomolecule-collection media are disposed sequentially in relation to eachother as first and second media.
 7. A molecule separator device asclaimed in claim 6 wherein the at least two molecule-collection mediaare disposed in any order in relation to each other.
 8. A moleculeseparator device as claimed in claim 6 wherein the at least twomolecule-collection media are disposed sequentially axially in relationto each other.
 9. A molecule separator device as claimed in claim 3wherein the first molecule-collection medium is at least an ion-exchangemembrane, a hydrophobic membrane, an affinity membrane or a combinationof any two or all three thereof.
 10. A molecule separator device asclaimed in claim 9 wherein the first molecule-collection medium isfabricated of material selected from the group consisting of nylon,polycarbonate, polyethersulfone, polysulfone, glass fiber,polypropylene, cellulose acetate, regenerated cellulose, and mixedesters of cellulose.
 11. A molecule separator device as claimed in claim10 wherein the second molecule-collection medium is fabricated ofmaterial selected from the group consisting of nylon, polycarbonate,polyethersulfone, polysulfone, glass fiber, polypropylene, celluloseacetate, regenerated cellulose, and mixed esters of cellulose.
 12. Amolecule separator device as claimed in claim 5 wherein the secondmolecule-collection medium is fabricated of material selected from thegroup consisting of nylon, polycarbonate, polyethersulfone, polysulfone,glass fiber, polypropylene, cellulose acetate, regenerated cellulose,and mixed esters of cellulose.
 13. A molecule separator device asclaimed in claim 12 wherein the second molecule-collection medium isanisotropic.
 14. A molecule separator device as claimed in claim 1wherein the housing is generally cylindrical for operational acceptancewithin a generally cylindrical fixed-angle or swinging-bucket chamber ofa centrifuge head.
 15. A molecule separator device as claimed in claim14 wherein the housing comprises a plurality of liquid-tight,releasably-connected compartments in communication with each other. 16.A molecule separator device as claimed in claim 1 wherein the housingadditionally comprises a releasably-connected cap with an aperture therethrough for receiving a fluid or gas pressurization nozzle.
 17. Aprocess for isolating from a solution thereof molecules having at leasttwo separable properties, the process comprising: a) placing thesolution into a compartment situated above a first-property specificfilter medium of a molecule separator device; b) driving the solutionforward through the first-property specific filter medium for capturingon said first-property specific filter medium molecules possessing thefirst property; and c) releasing the molecules from the first-propertyspecific filter medium and driving said molecules backward to asecond-property specific filter medium for capturing on saidsecond-property specific filter medium molecules possessing both thefirst and second properties.
 18. A process as claimed in claim 17wherein the first property is ionic and/or hydrophobic and/or affinityattraction.
 19. A process as claimed in claim 18 wherein the secondproperty is a molecular weight property from about 5×10² to about 3×10⁶Daltons.
 20. A process as claimed in claim 17 wherein the secondproperty is a molecular weight property from about 5×10² to about 3×10⁶Daltons.
 21. A process as claimed in claim 17 wherein the molecules aredriven by centrifugal force.